CLIP-Vista Green

CLIP-Vista Green is a green fluorescent substrate that can be used to label CLIP-tag™ fusion proteins (in cell lysates or purified proteins) for detection by SDS-PAGE. This substrate (BC-Vista Green) is based on fluorescein and is optimized for excitation with the 488 nm laser excitation line in a laser based gel scanner. It can also be excited using 360 nm light from a standard UV-transilluminator. It has an excitation maximum at 500 nm and emission maxima at 524 nm. This package includes 50 nmol of CLIP-Vista Green substrate, sufficient to label one hundred 20 µl samples containing CLIP-tag fusion protein for in-gel detection.

The CLIP-tag™ protein labeling system enables the specific, covalent attachment of virtually any molecule to a protein of interest. CLIP-tag is a protein tag based on human O6-alkylguanine-DNA-alkyltransferase (hAGT). CLIP-tag substrates are derivatives of benzylcytosine (BC). In the labeling reaction, the substituted benzyl group of the substrate is covalently attached to the reactive cysteine of CLIP-tag forming a stable thioether bond. Although CLIP-tag is based on the same protein as SNAP-tag, the benzylcytosine substrates form a separate class of substrates, different from the benzylcytosine substrates form a separate class of substrates, different from the benzylguanine substrates recognized by SNAP-tag. CLIP-tag and SNAP-tag® can be used for orthogonal and complementary labeling of two proteins simultaneously in the same cells.

There are two steps to using this system: subcloning and expression of the protein of interest as a CLIP-tag fusion, and labeling of the fusion with the CLIP-tag substrate of choice. Expression of CLIP-tag fusion proteins is described in the documentation supplied with CLIP-tag plasmids. The labeling of the fusion proteins with the CLIP-tag substrate for detection by SDS-PAGE is described in this document.Figure 1:
Excitation (dotted line) and emission spectra of CLIP-Vista Green coupled to CLIP-tag in buffer at pH 7.5
Figure 2:
Typical SDS-PAGE of CLIP-Vista Green labeled proteins visualized using a gel scanner. The gel was imaged with a Typhoon 9400 imager at 300V PMT with the 488/526 nm excitation/emission filter set.
Lane 1 fluorescent MW marker
Lanes 2–5 CLIP-His (22 kDa) 50, 150, 300, and 600 ngFigure 3: Structure of CLIP-Vista Green (MW 588.6 g/mol)

Excitation

Emission

Storage Temperature

Storage: Store substrate at -20°C. After dissolving CLIP-Vista Green store it in aliquots at -20°C in the dark for up to three months. Once an aliquot is thawed it can be stored at 4°C in the dark for up to two weeks. Make sure substrate, which may precipitate during freezing, is completely dissolved before use.

With proper storage at -20°C, the substrate should be stable for at least three years dry, or three months dissolved in DMSO.

The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at [email protected] or fill out the Technical Support Form for appropriate document.)

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at [email protected] or fill out the Technical Support Form for appropriate document.)

This product is covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB).

While NEB develops and validates its products for various applications, the use of this product may require the buyer to obtain additional third party intellectual property rights for certain applications.

For more information about commercial rights, please contact NEB's Global Business Development team at [email protected].

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

Licenses

Notice To Buyer/User: The Buyer/User has a non-exclusive license to use this system or any component thereof for RESEARCH AND DEVELOPMENT PURPOSES ONLY. Commercial use of this system or any components thereof requires a license from New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938.

Cellular Imaging and Analysis (i.e., SNAP and CLIP products)

Notice to Buyer/User: The Buyer/User has a non-exclusive license to use this system or any component thereof for RESEARCH AND DEVELOPMENT ONLY. Commercial use of this system or any components thereof requires a license from New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938.

The products and/or their use may be covered by one or more of the following patents and patent applications:

These patents and patent applications are owned by Covalys, or owned by the Ecole Polytechnique Fédérale de Lausanne (EPFL) and exclusively licensed to Covalys and NEB.

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